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Interleukin

Posted by Karen on March 19th, 2015  ⟩  0 comments

The difference between growth factors and transcription factors might be a nonissue for many researchers, but after Google searching, I have discovered that the question does exist within our field.

My introduction to transcription factors came from my undergrad developmental biology class. The problem is that my professor classified both growth factors and transcription factors, all as transcription factors. And since this was my first time learning about it, there was no reason to question his lecture. I must also admit that my textbook either did not do a great job defining the difference, or I just wasn’t looking for it. For me, things like fibroblast growth factors all fell under transcription factors. I didn’t consider them to be synonymous; instead, I thought growth factors were simply a subcategory.

It wasn’t until coming to GoldBio that I began to question the possibility of a difference between the two. I’m not alone in this revelation, which is why I think it’s important to highlight the defining differences.

Transcription Factors: Transcription factors are molecules that can bind either directly or indirectly to a DNA sequence and regulate the transcription of a particular gene or set of genes. They function in concert with other proteins, either blocking or promoting RNA polymerase. Transcription factors also have at least one, but sometimes more, DNA binding domains that allows them to attach to specific sequences or to DNA near the gene being regulated.

Growth Factors: Growth factors are generally molecules that are secreted and interact with other molecules or specified receptors to influence cellular behavior, including cell differentiation, healing and cell proliferation.

Key Difference: The major difference is that transcription factors bind to DNA while growth factors do not. Instead of binding to DNA, growth factors interact with other cellular molecules.

The Relationship: Part of the natural confusion comes from the relationships that exist between the two. As you learn about different signaling pathways, the primary goal when you’re first introduced to the topic is to understand the interaction between molecules and proteins within a particular cascade. Once you wrap your head around that, it’s easy to overlook some of the physical and chemical differences that exist between each player.

In a given pathway, a growth factor is secreted by a cell. It then binds to its cell surface receptor, and that interaction catalyzes a chain of actions within the cell. Binding leads to signal transduction until finally a transcription factor is activated, or a cell’s growth behavior is influenced.

Naturally, that means that all the interactions must be especially fine-tuned for optimal performance. This is why we see horrible developmental disorders or cancers when a particular protein is mutated. Blocking part of a pathway can lead to a disruption in the whole process.

To further clarify it, here is a list of known mouse transcription factors and here is a list of GoldBio growth factors, which are used in a wide variety of developmental research and cancer research.

So now the confusion is dispelled. Perhaps you didn’t even know you were confused.


    
              Karen Martin
GoldBio Marketing Coordinator


"To understand the universe is to understand math." My 8th grade
math teacher's quote meant nothing to me at the time. Then came
college, and the revelation that the adults in my past were right all
along. But since math feels less tangible, I fell for biology and have
found pure happiness behind my desk at GoldBio, learning, writing
and loving everything science. 



Category Code: 88221 79108 88253

Posted by Chris on October 10th, 2013  ⟩  0 comments

What if cases of delirium or instances of delusions were only a symptom of an imbalance of inflammatory growth factors in your brain? What if the root cause for debilitating mental diseases, such as Alzheimer’s, were really grounded in the never-ending tug and pull balancing act between cytokines and chemokines, their receptors and their inhibitors that goes on inside each of us every day? What if the whole of “getting older” (the joint pain, the memory lapses, etc.) was just a side effect of the messed up secretions of interleukins, fibroblast growth factors (FGFs), and other growth factors that we unknowingly depend on without even knowing what for? Those are some of the questions which motivated Dunja Westhoff and colleagues from the University of Amsterdam to try to find out.

esthoff and her group decided to look into the pre-operative expression pattern of cytokines in elderly patients who were admitted for hip fracture. A large percentage of elderly patients with hip fractures suffer from post-operative delirium, which seems to make sense since both the fracture and the surgery lead to systemic responses. Of course, discovering the subtleties of cytokine expression in elderly patients is no walk in the park. They were unable to detect 5 cytokines at all and 21 other cytokines were only detectable in a minority of the patients…leaving just 22 growth factors (including FGF2, IL1B, IL2, IL3, IL4, IL6, TNF-α, and EGF) that could be measured across both patients suffering from delirium and those who did not.

They eventually focused on 4 cytokines, three that had significantly lower levels (Flt-3L, IL1RA, and IL6) and one that was significantly higher (IP-10) in post-operative delirious patients. IL1RA (also listed as IL1RN) is a natural inhibitor of IL1A and IL1B, which are both pro-inflammatory cytokines in the brain. Subsequently, the reduction of IL1RA would lead to higher inflammation in the brain. Decreased levels of IL1RA have also been previously observed in patients suffering from either Alzheimer’s or rheumatoid arthritis.

This is a small study, and knowingly limited in its findings. But there seems to be a spark of truth amongst the clutter of results. Westhoff’s results seem to agree that there is a neuroinflammatory response effect which may be at least partially responsible for cases of delirium. Consequently, this may become an exciting avenue of clinical study and research. More research is necessary in order to prove if the effect is due to heightened proinflammatory responses or reduced anti-inflammatory responses. As usual, we have so much yet to learn. But, if true, this should lead to some exciting breakthroughs!

 
 

Westhoff, D., et al. (2013). "Preoperative cerebrospinal fluid cytokine levels and the risk of postoperative delirium in elderly hip fracture patients." J Neuroinflammation 10(1): 122.

Category Code: 88241

Posted by Chris on October 3rd, 2013  ⟩  0 comments

Necrosis is a very bad thing. I used to think that all cell death was bad, but that was before I began to understand the difference between apoptosis and necrosis. In its most basic form: Apoptosis = good cell death and Necrosis = bad cell death. That may be a little too simple, but it was the beginning of my understanding of some of the intricacies of cellular response to damage, disease or stress.

There has been a lot of research over the last 10 years focusing on cell stress and cell death and the molecular pathway differences between the two types of cell death. In apoptosis, the programmed death of a cell, cells naturally expire and produce apoptotic bodies which can be properly engulfed by phagocytes and removed before they can cause damage. The process of apoptosis is critical in embryonic development for processes like separating our fingers into individual digits and the overexpression or underexpression of apoptosis can cause problems like atrophy or cancer.

Necrosis, on the other hand, is the completely unplanned death of a cell, or at least unplanned by the cell. There are lots of causes for necrosis; including physical injury, infection, or toxicity. Cells that undergo necrosis do not elicit the proper signal pathways for the phagocytes. Instead, they uncontrollably release their contents into the intracellular space, causing inflammation or spreading the cause for the necrosis into surrounding cells. The result is a cascading effect of cell death that results in things like gangrene.

As scientists have begun to understand this process, one of the things they discovered was a wide array of damage associated molecular pattern molecules, or more easily called DAMPs. DAMP molecules are commonly released in the process of necrosis, but are prevented from release during apoptosis. IL1A is one such DAMP molecule that is responsible for activating an immunity response. The immunity response from DAMPs causes sterile inflammatory response which has been shown to be a factor in many diseases including atherosclerosis, ischemia reperfusion injury and Alzheimer's. Increased IL1 activity has also been associated with diabetes, rheumatoid arthritis, gout, and psoriasis.

IL1R1Since IL1A is universally expressed, it is believed to act as a universal DAMP molecule, but the actual process has still be unclear. While it was known that the pro-IL1A (p33) is processed to the mature IL1A (p17) by calpain, it was not known what the consequences were for that cleavage. Yue Zheng et al. describe in a recent paper that the cleavage of p33 is essential for IL1A and increases its affinity for IL1R1 (Receptor 1). What they found was that IL1A is primarily bound by IL1R2 (Receptor 2), which protects it from cleavage and prevents its activity. In fact, without IL1R2, the most significant DAMP in necrotic cells is IL1A. They showed that IL1R1 rich tissue (e.g. liver) activated the immune response regardless of IL1A cleavage, whereas IL1R1 deficient tissue (e.g. kidney) that cannot cleave p33 did not respond to IL1A at all.

There are other DAMP molecules, of course, and IL1A isn’t the only molecule that can drive inflammation, but Zheng points out that the additional activation of IL1A might just act as the tipping point of the inflammatory response that drives it toward adaptive immunity. Zheng has found that IL1R2 controls the release of Il1A through the activation of caspase-1, which processes IL1R2 and releases IL1A for binding to IL1R1 and cleavage by calpain.

Fundementally, this paper presents a small, but meaningful step toward understanding the roles of these molecules play in situations like graft rejection or chronic diseases like Alzheimer's and atherosclerosis. And ultimately, we can hope that this knowledge will help doctors work around these types of problems in the future.

 
 

Zheng, Y., Humphry, M., Maguire, J. J., Bennett, M. R., & Clarke, M. C. (2013). Intracellular Interleukin-1 Receptor 2 Binding Prevents Cleavage and Activity of Interleukin-1α, Controlling Necrosis-Induced Sterile Inflammation. Immunity. 38, 285–295

Category Code: 88221 88241

Posted by Chris on June 20th, 2013  ⟩  0 comments

If you’re anything like me (geek that I am), every new technological device tends to get your blood pumping and invokes an involuntary reflex to reach for your wallet. That’s even truer for the ever-popular “i”-products which tend to grab our collective-geek attention even faster with every new device. Now there are some clever scientists from the University of Bonn in Germany who have developed a new reporter system utilizing Gaussia luciferase, the “iGLuc”!

While researching the inflammasome process, and specifically IL-1β, a primary target of caspase-1, Bartok et al. hit a frustrating road block. Inflammasomes are large, multiprotein oligomers that are intregal parts of the immune response system. They are a platform which supports an inflammatory cascade after sensing damage-associated molecular patterns. Caspase-1 is an enzyme that’s utilized by the inflammasome cascade in order to proteolytically cleave specific proteins (such as IL-1β precursor) into active, mature peptides. Once cleaved, IL-1β can finally bind to its receptor in order to induce a variety of cellular responses, such as pyroptosis; a form of programmed cell death that is in response to inflammation.

Bartok was looking for a better way to analyze IL-1β. ELISA techniques were not sensitive enough to distinguish between the IL-1β precursors and mature IL-1β, and Western blotting was too time consuming and useless for high throughput analysis. So, instead they devised a fusion protein of pro-IL-1β and GLuc (Gaussia Luciferase) and called it iGLuc! Unexpectedly, they first saw virtually no luciferase signal from the fusion, even though they were seeing high expression levels of luciferase in the system. But they discovered that pro-IL-1β tends to form a protein aggregate which acts to restrict the release of the signaling C-terminal portion of GLuc. But with the simple addition of caspase-1, pro-IL-1β was cleaved and a corresponding bioluminescent signal could be measured.

The resulting process seems to make for an excellent reporter assay for inflammasome activity! Bartok tested the system both in vitro and in vivo and the system showed good sensitivity and specificity as well as a great signal to noise ratio. The system also shows a lot of promise that it can be further applied to other proteases as well! So, if you’re in the field of inflammasomes (or if you have to own every new device), be sure to keep an “i” out for the iGLuc system. It may become the next, best geeky thing on the technological front! You can find their complete article here.

iGLuc in vivo images

Category Code: 88221 88241

Posted by Chris on April 4th, 2013  ⟩  0 comments

The interleukin family of cytokines is one of the largest and most studied of all the growth factors. Their roles in disease, the immune system, and immune deficiency have made them superstars of cancer research and AIDS/HIV research, not to mention as possible, critical links in such diverse problems such as heart disease, neurological disorders like Alzheimer’s, arthritis and even Crohn’s disease. Interleukins are involved in processes of cell activation, cell differentiation, proliferation, and cell-to-cell interactions. The expression of interleukins is usually strictly regulated, i.e., the factors are often not secreted constitutively. They are most often synthesized after cell activation as a consequence of a physiological or non-physiological stimulus. There are also some interleukins which are autoregulatory and regulate their own synthesis or the expression of their own receptors.

Gold Bio is excited to now offer three interleukins (IL2, IL3 and IL4), recombinant from both human and murine sequence for your research needs! IL2 was the first of the interleukin family to be identified and characterized in the early 1980’s, though the existence of this family of growth factors was known for a few decades before that. IL2 is necessary for the growth, proliferation, and differentiation of T cells to become 'effector' T cells. IL2 has been shown to be similar to IL15, but IL2 is instrumental in adaptive immunity and the development of the immunological memory, playing an important role in both regulatory T cells (Treg) development and function, whereas IL15 is more important in maintaining a highly specific T cell response.

IL3 is a popular cytokine in use for a variety of cell cultures (i.e. mast cells or basophils) providing the cytokinetic connection between the immune and hematopoietic systems. IL3 is capable of inducing the growth and differentiation of multi-potential hematopoietic stem cells, neutrophils, eosinophils, megakaryocytes, macrophages, lymphoid and erythroid cells. Haig, et al. recently showed a synergistic affect between IL3 and another growth factor, KITLG (sometimes called SCF or Stem Cell Factor), on both bone marrow-derived mast cells (BMMC) and serosal/connective-tissue mast cells (CTMC).

IL4 is most closely associated with IL13 and induces native T helper (Th0) cells to become Th2 cells (which then produce more IL4). They are often produced during allergic responses and promote allergic inflammation by activation signal tranducers. IL4 actions are often “neutralized” by Inferon-gamma (IFN-ɣ), which is made by the Th1 cells. Gilbert, et al., showed that IL4 (as well as IL1) are involved in the response of annulus fibrosus (AF) cells derived from nondegenerative tissue to cyclic tensile strain.

If you have any questions about interleukins or any of our other available growth factors, you can contact us at: techsupport@goldbio.com!

 
 

Mahmud, Shawn A., Luke S. Manlove, and Michael A. Farrar. "Interleukin-2 and STAT5 in regulatory T cell development and function." JAK-STAT 2.1 (2013): 0-1.

Haig, David M., et al. "Effects of stem cell factor (kit-ligand) and interleukin-3 on the growth and serine proteinase expression of rat bone-marrow-derived or serosal mast cells." Blood 83.1 (1994): 72-83.

Reddy, E. Premkumar, et al. "IL-3 signaling and the role of Src kinases, JAKs and STATs: a covert liaison unveiled." Oncogene 19.21 (2000): 2532-2547.

Gilbert, Hamish TJ, et al. "The involvement of interleukin-1 and interleukin-4 in the response of human annulus fibrosus cells to cyclic tensile strain: an altered mechanotransduction pathway with degeneration." Arthritis research & therapy 13.1 (2011): R8.

Category Code: 88221 79108